Greatest Homology Research Articles

Isolation and expression of a cDNA encoding the precursor for a novel member (ACADSB) of the acyl-CoA dehydrogenase gene family.

The acyl-CoA dehydrogenases (ACDs) are a family of mitochondrial enzymes that oxidize straight chain or branched chain acyl-CoAs in the metabolism of fatty acids or branched chain amino acids. Deficiencies in members of this gene family are important causes of human disease. A cDNA encoding the human precursor for a novel member (gene symbol ACADSB) of the ACD gene family has been isolated and characterized. The open reading frame of 1.3 kb encodes a precursor protein of 431 amino acids, which is processed in vitro to yield a mature protein of 399 amino acids. The cDNA has significant sequence similarity to other members of the acyl-CoA dehydrogenase family, with the greatest homology (38%) to the short chain acyl-CoA dehydrogenase. The cDNA was expressed in eukaryotic (COS) and prokaryotic (Escherichia coli) cells, producing a protein of the expected size, with activity toward the short branched chain acyl-CoA derivatives ((S)-2-methylbutyryl-CoA, isobutyryl-CoA, and 2-methylhexanoyl-CoA), as well as toward the short straight chain acyl-CoAs (butyryl-CoA and hexanoyl-CoA).

Identification of the major brown shrimp (Penaeus aztecus) allergen as the muscle protein tropomyosin.

Shrimp, a major seafood allergen, was investigated as a model food allergen. Extracts from both shrimp (Penaeus aztecus) meat and cooking fluid contain a substantial and similar amount of allergenic activity. A 36-kD allergen, demonstrated in both extracts by SDS-PAGE/Western blot analysis, reacted with 28/34 (82%) sera from shrimp-sensitive, skin test and RAST-positive, individuals. This allergen, named Pen a I, was isolated by SDS-PAGE; its amino acid composition was rich in aspartic and glutamic acids. A 21-residue peptide, obtained from endoproteinase Lys-C digested Pen a I by high-performance liquid chromatography, demonstrated significant homology (60-87%) with the muscle protein tropomyosin from various species and origins. The greatest homology (87%) was noted with tropomyosin of the fruit fly (Drosophila melanogaster) reflecting the phylogenic relationship between these two arthropods. These studies demonstrate that tropomyosin is the major shrimp allergen. Although the amino acid sequence of this shrimp muscle protein shares considerable homology with tropomyosins of other species including man, significant differences remain in allergenic activity.

Molecular cloning of cDNA encoding the precursor for hamster hypothalamic growth hormone-releasing factor

The structure of rat and mouse growth hormone-releasing factor (GRF) peptide and precursor shows considerable divergence from that of the human counterpart and also within rodents themselves. To study such structural divergence in another rodent, we cloned a cDNA encoding the GRF precursor from golden hamster. The hamster GRF (haGRF) cDNA clone had an open-reading frame that predicts a haGRF precursor protein with 107 amino acids. The haGRF precursor bore greater overall homology (82%) to the human than the same rodent homologue (58-64%) and contained two processing sites identical to the human sequence that would generate mature haGRF peptide. Furthermore, the haGRF peptide, like human but unlike rat or mouse GRF, consisted of 44 amino acids and also had greater homology to the human (89%) than the rodent sequence (64-75%), conserving a Tyr residue at the N-terminus and an amidated Leu residue at the C-terminus. Thus, both haGRF precursor and peptide are structurally more related to those of human than of other rodents, suggesting that rodent GRF precursor diverged from the human sequence at differential rates within the species.

Expression, purification, and characterization of the G protein-coupled receptor kinase GRK5.

G protein-coupled receptor kinases (GRKs) such as rhodopsin kinase and the beta-adrenergic receptor kinase (beta ARK) play an important role in agonist-specific phosphorylation and desensitization of G protein-coupled receptors. GRK5 is a recently identified member of the GRK family that has greater homology with rhodopsin kinase than with beta ARK. To further characterize the activity of GRK5, it has been overexpressed in Sf9 insect cells and purified by successive chromatography on S-Sepharose and Mono S columns. GRK5 phosphorylates the beta 2-adrenergic receptor (beta 2AR), m2 muscarinic cholinergic receptor, and rhodopsin in an agonist-dependent manner to maximal stoichiometries of approximately 2.5, 1.5, and 1 mol of phosphate/mol of receptor, respectively, with Km values of approximately 0.5 microM for the beta 2AR, approximately 16 microM for rhodopsin, and approximately 24 microM for ATP. Peptide phosphorylation studies suggest that in contrast to beta ARK and rhodopsin kinase, GRK5 preferentially phosphorylates on nonacidic peptides with a Km of approximately 1.5 mM. Heparin and dextran sulfate were found to be potent inhibitors of GRK5 with IC50 values of approximately 1 nM, thereby being at least 150-fold more potent on GRK5 than on beta ARK. GRK5 can also be activated by polycations, with 10 microM polylysine promoting an approximately 2.6-fold activation. Overall, these studies demonstrate that GRK5 has unique properties that distinguish it from other members of the GRK family and that likely play an important role in modulating its mechanism of action.

Sequence and expression of endogenous S-locus glycoprotein genes in self-compatible Brassica napus

Brassica napus is an amphidiploid plant which is self-compatible even though it is derived from hybridisation of the self-incompatible species B. oleracea and B. campestris. Experiments were undertaken to establish if S-locus glycoprotein (SLG) genes exist in B. napus and whether these are expressed as in self-incompatible Brassica species. Two different stigma-specific cDNA sequences homologous to SLG genes were obtained from the B. napus cultivar Westar. One of these sequences, SLGWS1, displayed highest homology to class I SLG alleles, whereas the other, SLGWS2, showed greatest homology to class II SLG genes. Both were expressed at high levels in Westar stigmas following a developmental pattern typical of SLG genes in the self-incompatible diploids. We infer that they represent the endogenous SLG genes at the two homologous S-loci. The occurrence of normally expressed SLG genes and its relevance to the self-compatible phenotype of B. napus is discussed.

New isoforms of multifunctional calcium/calmodulin-dependent protein kinase II

Calcium/calmodulin dependent protein kinase II (CaM kinase II) seems to act as an important regulator of intracellular signal transmission. Four subtypes, termed α to δ, have been cloned; some of them can exist as different splicing variants. All these isoforms share a great overall homology, and they contain 3 areas of low homology. We have identified 5 new variants of subtype delta so that the total number of different isoforms now adds up to 12. These variants are probably a result of different splicing and show several deletions in regard to subtype delta. The deletion sites do exactly match regions of low homology between the subtypes. This suggests a functional division of the CaM kinase II molecule into homologous and variable domains. The homologous domains are highly conserved. Therefore, it might be the case that the constitution of the variable domains is more significant for a certain isoform than its belonging to one of the 4 subtypes α to δ.

Can a protein influence the fate of its own coding sequence?: the amino- and carboxyl-terminal regions of H1 histone.

The amino- and carboxyl-terminal regions of H1 histone are afforded an opportunity to bind to their own coding segments. While such binding is of no consequence to the type of H1 histone that is expressed only in terminally differentiated cells (here referred to as H5), the same binding occurring in H1 of proliferating cells might affect the course of evolution of its amino- and carboxyl-terminal regions. With the above in mind, amino acid sequences of H5 from three species were compared with those of H1 from five species. The spherical core was more conserved in H1 than in H5, whereas the amino- and carboxyl-terminal regions of both were hypervariable. A distinct evolutionary pattern, however, was observed only in these regions of H1 but not of H5. Tandem repeats of short palindromes often generated nearly identical long palindromes in distantly related species. Thus, as far as the 37-residue-long amino-terminal region was concerned, the greatest homology was between the rat and human rather than between the rat and the mouse. With regard to the last 25 residues in the carboxyl-terminal region, the greatest homology was between the chicken and the rainbow trout; the next was between the mouse and the trout. Successions of long palindromes populating the amino- and carboxyl-terminal regions of H1 might be the direct consequence of the autologous binding between peptides and their coding sequence.

A Campylobacter jejuni homolog of the LcrD/FlbF family of proteins is necessary for flagellar biogenesis

A Campylobacter jejuni homolog of the lcrD/flbF family of genes was cloned and sequenced. The nucleotide sequence of the gene, called flbA, predicted a protein of 78,864 Da, with significant homology to a group of related proteins including the Yersinia pestis LcrD, Salmonella typhimurium InvA, and Caulobacter crescentus FlbF proteins. The greatest homology was seen with the C. crescentus FlbF protein, with an overall amino acid sequence homology of 57%. An insertion mutation in the C. jejuni 81-176 flbA gene was constructed. The resultant strain did not synthesize flagellin and was nonmotile.

Molecular cloning and expression of rabbit pancreatic cholesterol esterase

Rabbit pancreatic cholesterol esterase (CEase, carboxyl ester lipase, EC 3.1.1.3) has been cloned from a λgt11 library of adult rabbit pancreatic cDNA. The open reading frame consists of 1788 nucleotides which encodes 576 amino acids of the functional protein and a 20 amino acid leader peptide. When compared to other species, the greatest homology is observed between residues 82–248 with little or no homology at the C-terminal end where proline-glutamate-serine-threonine (PEST) segments are a characteristic feature of the human CEase. Rabbit CEase (RCEase) retains the active-site serine (gxsxg), the active-site histidine and the tentative heparin binding site (KKRCLQ) at similar positions in comparison to pancreatic CEases of other species. When rabbit CEase cDNA is expressed in monkey kidney (COS-7) cells, enzymatic hydrolytic activity is detected in the growth medium as is a 67 kDa protein by Western blotting with polyclonal anti-CEase antibody. Northern blot analysis shows two mRNA (2.2 and 3.2 kb) species.

Identification of nuclear factors which interact with the 5′ flanking region of the EF-1αO gene in Xenopus laevis

The EF-1αO gene of Xenopus laevis is a stage-specific gene, being transcribed in oogonia and oocytes, but not in postmeiotic germ cells and terminally differentiated cells. We found that two trans-acting factors from oocyte nuclear extract are able to interact with a DNA sequence in the 5′-upstream region of the EF-1αO gene. Methylation interference experiments suggested that the two factors recognised the same DNA element. Gel retardation assays indicated that part of the protein binding site could be confined to a 21 bp sequence, located between −51 and −72, relative to the cap site. Interestingly, this region shares great homology to a negative regulatory segment in the promoter of the TFIIIA gene, another developmentally regulated gene.

Cloning, sequence and expression of a β-tubulin-encoding gene in the homobasidiomycete Schizophyllum commune

The β-tubulin (βTub)-encoding gene ( tub-2) of Schizophyllum commune is the first tubulin gene isolated, cloned and sequenced from higher filamentous fungi (homobasidiomycetes). The S. commune tub-2 gene is organized into nine exons and eight introns. The introns vary from 48 to 107 nt in length, and are distributed throughout the gene. The tub-2 exons code for a protein of 445 amino acids (aa), which shows great homology with βTubs of filamentous ascomycetes, plants, and animals, but less homology with yeasts. The codon usage of tub-2 from S. commune is biased, as it is in most βTub-encoding genes of filamentous fungi. The S. commune βTub shows a conserved aa sequence in the C-terminal domain, which is suggested to interact with microtubule-associated proteins in animals. In contrast, the S. commune βTub deviates from most known βTubs by having a Cys 165 residue, which might be significant for the insensitivity of S. commune haploid strains to the antimicrotubule drug, benomyl. In tub-2 of different haploid strains, sequence polymorphisms occur in the 5' and 3' flanking regions. The expression of tub-2 is high in young mycelium, which has a high number of extending apical cells, but decreases with the aging of the mycelium. No significant difference in the hybridization signal intensity for the tub-2 transcripts was recorded either during intercellular nuclear migration at early mating, or in mycelia with a mutation in the B mating-type gene. The weak signal obtained with the 3' end of tub-2 as a probe in Southern hybridizations, under conditions of low stringency, warrants attention to the possibility that S. commune might have another βTub-encoding gene highly divergent from tub-2.

Ugandan HIV-1 V3 loop sequences closely related to the U.S./European consensus

The third variable (V3) loop of the human immunodeficiency virus type 1 (HIV-1) envelope protein is an important determinant for virus neutralization and cell tropism. V3 loop sequences from uncultured lymphocytes obtained in 1990 from 22 Ugandan HIV-1-infected patients could, with the exception of two patients' sequences, be divided into two groups (A and B) on the basis the V3 loop size and sequence. The V3 loop consensus sequences from both groups showed a high degree of homology to a U.S./European consensus, a characteristic also reflected by the results of peptide serology. In the case of group B the difference in sequence was only five amino acids. In contrast, the V3-flanking regions for both groups showed greater homology to an earlier (1986/1987) Ugandan consensus. The discovery of these two new Ugandan V3 loop genotypes, which are closely related to the U.S./European consensus, has implications for the understanding of the evolution of HIV-1 and for the future design of a vaccine for use in Africa.

Isolation and sequencing of the cDNA coding for spinach 10-formyltetrahydrofolate synthetase. Comparisons with the yeast, mammalian, and bacterial proteins.

The one-carbon metabolism enzymes 10-formyltetrahydrofolate synthetase (EC 6.3.4.3), 5,10-methenyltetrahydrofolate cyclohydrolase (EC 3.5.4.9), and 5,10-methylenetetrahydrofolate dehydrogenase (EC 1.5.1.5) can be found on a single trifunctional protein in the eukaryotes examined. The one exception is in spinach leaves where 10-formyltetrahydrofolate synthetase is monofunctional (Nour, J. M., and Rabinowitz, J. C. (1991) J. Biol. Chem. 266, 18363-18369). In the prokaryotes examined, 10-formyltetrahydrofolate synthetase is either absent or is monofunctional. A cDNA clone encoding spinach leaf 10-formyltetrahydrofolate synthetase was isolated through the use of antibodies to the purified enzyme. This clone had an open reading frame of 1914 base pairs and encoded for a protein containing 636 amino acids with a calculated M(r) of 67,727. The percentage identity between spinach 10-formyltetrahydrofolate synthetase and the synthetase domains in the four trifunctional eukaryotic enzymes and the two monofunctional prokaryotic enzymes that have been cloned and sequenced was: 64.9% human, 63.8% rat, 55.6% yeast cytoplasm, 53.8% yeast mitochondria, 47.8% Clostridium acidi-urici, and 47.9% Clostridium thermoaceticum. Clearly the spinach monofunctional protein had greatest homology with the mammalian proteins. The spinach protein is longer than the two other monofunctional prokaryotic proteins. Possible reasons for this are presented. The codon usage and the putative translation initiation sites are examined and compared with other spinach proteins.

Methanogenesis from acetate: a comparison of the acetate metabolism inMethanothrix soehngeniiandMethanosarcinaspp.

Abstract Acetate is the precursor of approximately two-thirds of the methane produced in anaerobic bioreactors. Only two genera of methanogenic archae are known to use acetate as sole energy source: Methanosarcina and Methanothrix. Methanosarcina appears to be a generalist with a high growth rate, but low affinity for acetate. Methanothrix is a specialist having a high affinity for acetate, but low growth rate. Methanothrix shows a much lower minimum threshold for acetate utilization (7–70 μM) than Methanosarcina (0.2–1.2 mM). This is consistent with the evidence that Methanothrix is found in environments with low acetate concentrations. The acetate degradation by acetotrophic methanogens starts with an activation of acetate to acetyl-coenzyme A. In Methanosarcina spp. this activation is catalysed by an acetate kinase/phosphotransacetylase system at the expense of one ATP. Acetyl-coenzyme A synthetase activates acetate in Methanothrix, with concomitant hydrolysis of one ATP to AMP and PPi. Both enzyme systems have been purified and comparison of the kinetic properties confirmed the hypothesis that low acetate concentrations favour Methanothrix. The gene encoding for acetyl-CoA synthetase of Methanothrix was isolated from a genomic library and actively expressed in Escherichia coli. The deduced amino acid sequence showed homology to proteins with similar function and contained two putative ATP binding sites. The most characteristic and complex enzyme involved in the acetate degradation by acetotrophic methanogens is carbon monoxide dehydrogenase. The enzyme has been purified from both Methanothrix and Methanosarcina, and represents 5–10% of the soluble protein of these microorganisms. CO dehydrogenase is proposed to catalyse both the cleavage of acetyl-CoA in a methyl-, carbonyl- and CoA-moiety, and the oxidation of the carbonyl group to CO2. This multifunctional redox enzyme contains several iron, acid-labile sulfur and nickel atoms. These atoms are arranged into several paramagnetic complexes, which have been studied by EPR spectroscopy. The low spin recovery of the different paramagnetic centers makes statements about structure and functions difficult. There are good spectroscopic and genetic indications that the CO dehydrogenase of Methanothrix contains at least one ferrodoxin-like [4Fe-4S] cluster, which could play a role in the electron transfer of the CO oxidation. Further, in EPR spectra of concentrated samples of CO dehydrogenase from Methanothrix a very unusual signal was observed, which showed great similarity to putative [6Fe-6S] prismane clusters. The final step in the methanogenesis from acetate, the reduction of methyl-coenzyme M, is catalysed by methyl-coenzyme M methylreductase. The enzyme purified from Methanothrix and Methanosarcina showed great homology with the methyl-CoM methylreductase of other methanogenic archae, although the specific activity was rather low (60–125 nmol min−1 mg−1). The reduction of the heterodisulfide between coenzyme M and component B is proposed to be the common site for energy conservation in all methanogens. Acetoclastic methanogens, however, need additional sites of energy conservation to compensate for their high energy input in acetate activation. The oxidation of CO to CO2 could form one possible site. The partially membrane-associated pyrophosphatase of Methanothrix could form another site of energy conservation.

Methanogenesis from acetate: a comparison of the acetate metabolism in Methanothrix soehngenii and Methanosarcina spp.

Acetate is the precursor of approximately two-thirds of the methane produced in anaerobic bioreactors. Only two genera of methanogenic archae are known to use acetate as sole energy source: Methanosarcina and Methanothrix. Methanosarcina appears to be a generalist with a high growth rate, but low affinity for acetate. Methanothrix is a specialist having a high affinity for acetate, but low growth rate. Methanothrix shows a much lower minimum threshold for acetate utilization (7–70 μM) than Methanosarcina (0.2–1.2 mM). This is consistent with the evidence that Methanothrix is found in environments with low acetate concentrations. The acetate degradation by acetotrophic methanogens starts with an activation of acetate to acetyl-coenzyme A. In Methanosarcina spp. this activation is catalysed by an acetate kinase/phosphotransacetylase system at the expense of one ATP. Acetyl-coenzyme A synthetase activates acetate in Methanothrix, with concomitant hydrolysis of one ATP to AMP and PP i. Both enzyme systems have been purified and comparison of the kinetic properties confirmed the hypothesis that low acetate concentrations favour Methanothrix. The gene encoding for acetyl-CoA synthetase of Methanothrix was isolated from a genomic library and actively expressed in Escherichia coli. The deduced amino acid sequence showed homology to proteins with similar function and contained two putative ATP binding sites. The most characteristic and complex enzyme involved in the acetate degradation by acetotrophic methanogens is carbon monoxide dehydrogenase. The enzyme has been purified from both Methanothrix and Methanosarcina, and represents 5–10% of the soluble protein of these microorganisms. CO dehydrogenase is proposed to catalyse both the cleavage of acetyl-CoA in a methyl-, carbonyl- and CoA-moiety, and the oxidation of the carbonyl group to CO 2. This multifunctional redox enzyme contains several iron, acid-labile sulfur and nickel atoms. These atoms are arranged into several paramagnetic complexes, which have been studied by EPR spectroscopy. The low spin recovery of the different paramagnetic centers makes statements about structure and functions difficult. There are good spectroscopic and genetic indications that the CO dehydrogenase of Methanothrix contains at least one ferrodoxin-like [4Fe-4S] cluster, which could play a role in the electron transfer of the CO oxidation. Further, in EPR spectra of concentrated samples of CO dehydrogenase from Methanothrix a very unusual signal was observed, which showed great similarity to putative [6Fe-6S] prismane clusters. The final step in the methanogenesis from acetate, the reduction of methyl-coenzyme M, is catalysed by methyl-coenzyme M methylreductase. The enzyme purified from Methanothrix and Methanosarcina showed great homology with the methyl-CoM methylreductase of other methanogenic archae, although the specific activity was rather low (60–125 nmol min −1 mg −1). The reduction of the heterodisulfide between coenzyme M and component B is proposed to be the common site for energy conservation in all methanogens. Acetoclastic methanogens, however, need additional sites of energy conservation to compensate for their high energy input in acetate activation. The oxidation of CO to CO 2 could form one possible site. The partially membrane-associated pyrophosphatase of Methanothrix could form another site of energy conservation.

H chain V region sequences of three human monoclonal IgM with anti-myelin-associated glycoprotein activity.

The amino acid sequence corresponding to the V region H chain gene used by three monoclonal IgM directed to the myelin-associated glycoprotein (MAG) is presented. They all belonged to the VHIII variability subgroup, but each may well represent a new member of this family inasmuch as their homology with previously sequenced VHIII genes was less than 80%. Strikingly, there was no greater homology between the H chain V regions of the anti-MAG IgM. Partial amino acid sequence data indicated that these V regions were joined to as yet unidentified DH segments; however, two H chains used very similar DH, possibly indicating that this sequence was involved in the fine specificity of the IgM for MAG. All H chains included a JHIV region. These data, together with results obtained from the sequence of the three kappa L chains of the same IgM molecules (Mihaesco, E., H. Ayadi, N. Congy, M. C. Gendron, J. P. Roy, H. Heyermann, B. Frangione, and J. C. Brouet. 1989. J. Biol. Chem. 264:21481), indicate that the repertoire of VL and VH gene segments used by anti-MAG IgM is quite diverse, in contrast to previous structural data obtained for other human monoclonal IgM autoantibodies. Possibly, these differences reflect distinct pathogenesis.

Peptide recognition, T cell receptor usage and HLA restriction elements of human heat-shock protein (hsp) 60 and mycobacterial 65-kDa hsp-reactive T cell clones from rheumatoid synovial fluid.

A commonly held postulate regarding the etiology of rheumatoid arthritis (RA) is that of antigenic mimicry. Recent interest has focused on the mycobacterial 65-kDa heat-shock protein (hsp) as a putative causal agent. The 65-kDa hsp has over 40% sequence homology with the human hsp 60, and elevated synovial T cell responses to both antigens have been demonstrated in RA and juvenile rheumatoid arthritis patients. Such T cells should, therefore, be specific for shared epitopes on the two antigens. To investigate this, we screened synovial fluid mononuclear cells from two early RA patients with peptides of the 65-kDa hsp which have the greatest homology with the human hsp 60. We also raised a panel of T cell clones from one of the patients with the 65-kDa hsp. The synovial T cell population from both patients and one of the T cell clones recognized a peptide representing the amino-acid sequence 241-255. This clone also responded to the peptide of the equivalent human sequence, and was restricted by HLA-DQ. A second T cell clone recognized an adjacent epitope (amino acid sequence 251-265) which is also highly homologous with the human sequence, but this clone was restricted by HLA-DR. The clones utilized different V beta gene segments but the same D beta and J beta gene elements, and both exhibited specific cytotoxicity against autologous antigen-pulsed macrophages. Our findings, therefore, do not disagree with the postulate that autoimmune disease could possibly be triggered by bacterial epitopes with homology to self protein. However, it is also noted that there are alternative interpretations of this data.

Rainbow trout cytochrome P-450 c17 (17α-hydroxylase/17,20-lyase) cDNA cloning, enzymatic properties and temporal pattern of ovarian P-450 c17 mRNA expression during oogenesis

A cDNA clone encoding cytochrome P-450 c17 (17α-hydroxylase/17,20-lyase) was isolated from a rainbow trout ovarian follicle cDNA library. The cDNA contained an open reading frame of 1,542 nucleotides encoding a protein of 514 amino acid residues. The amino acid sequence of trout P-450 c17 shows a much greater homology with chicken P-450 c17 than with that of human, bovine and rat. The trout P-450 c17 expressed in non-steroidogenic mammalian COS-1 cells showed both 17α-hydroxylase and 17,20-lyase activities. The cDNA only hybridized to a single species or mRNA (2.4 kb) isolated from rainbow trout ovaries; the 2.4 kb transcripts were abundant in trout ovaries during the later stages of oogenesis.

Cloning of a cDNA encoding the ovine interleukin-2 receptor 55-kDa protein, CD25

A 1.3-kb cDNA that encodes the entire 825-bp coding region of ovine CD25, the interleukin-2 receptor 55-kDa protein, has been isolated. Comparison of the deduced amino acid sequence with CD25 proteins from other species shows the ovine sequence to have the greatest homology with that of the bovine species.

The complementary deoxyribonucleic acid sequence of guinea pig endometrial prorelaxin.

The nucleotide sequence of the relaxin gene transcript in the endometrium of the late pregnant guinea pig has been determined. The strategy used was a combination of polymerase chain reaction (PCR) with primers designed from the mRNA sequence of porcine preprorelaxin, rapid amplification of cDNA ends-PCR, and blunt end cloning in M13 mp18. With heterologous primers, a 226-basepair (bp) segment of the guinea pig relaxin gene sequence was obtained and was used to design a guinea pig-specific primer for use with the rapid amplification of cDNA ends-PCR method. The latter allowed completion of the sequence of 336 bp, with a 96-bp overlap. The sequence obtained shows greater homology at both the nucleotide and amino acid levels with porcine and human relaxins H1 and H2 than with rat relaxin, supporting the thesis that the guinea pig is not a rodent. The transcription of the guinea pig endometrial relaxin gene during pregnancy was confirmed by Northern analysis of guinea pig endometrial tissues with a species-specific cDNA probe. The endometrial relaxin gene is transcribed during pregnancy, but not in lactation, consistent with the observed immunostaining for relaxin.